The anti-tumor agent CCNU has the potential for alkylation of DNA, although the specifics have never been investigated. This project has extended into this area to determine the fate of cellular DNA after treatment of L1210 and rat glioma cells with CCNU both in vitro and in vivo. Our preliminary data indicates that exposure of cells to CCNU leads to the production of single strand breaks in DNA which can be demonstrated by analysis of the DNA or alkaline sucrose gradients or by hydroxylapatite chromatography. The latter, more sensitive method of analysis has detected the production of single strand breaks in DNA at CCNU concentrations well within the physiologically realistic range. In a study of this nature, the cell's ability to repair single strand breaks in DNA and the ability of the alkylating agent to produce these breaks are of equal importance. So far we have been unable to demonstrate repair of CCNU induced damage while knowing that, for example, gamma-irradiation damage is quickly repaired. Our data indicate that CCNU may act both to produce damage in DNA and to inhibit the cells repair process. Future work will examine this hypothesis both at the cellular level and in whole animal studies (Project Z01 CM 06903- 01 BCRC).